This invention relates to new peptide compounds and pharmaceutically acceptable salts thereof which are useful as medicament.
Some peptide compounds have been known as described in, for example, EP 0 394 989 A2, WO96/16981 and JP-A-10-81671.
This invention relates to new peptide compounds.
One object of this invention is to provide the new and useful peptide compounds and pharmaceutically acceptable salts thereof that possess a strong inhibitory activity on the production of nitric oxide (NO).
Another object of this invention is to provide a process for the preparation of the peptide compounds and salts thereof.
A further object of this invention is to provide a pharmaceutical composition comprising said peptide compound or a pharmaceutically acceptable salt thereof.
Still further object of this invention is to provide a use of said peptide compounds or pharmaceutically acceptable salts thereof as a medicament for prophylactic and therapeutic treatment of NO-mediated diseases including respiratory diseases such as adult respiratory distress syndrome (ARDS) and asthma; cardiovascular diseases such as cardiovascular ischemia, myocarditis, heart failure, hypotension and atherosclerosis; endocrine diseases such as diabetes (e.g., insulin-dependent diabetes mellitus, etc.), complications of diabetes mellitus (e.g., diabetic nephropathy, diabetic retinopathy, diabetic neuropathy, etc.) and gout; renal diseases such as glomerulonephritis and renal failure; gastrointestinal diseases such as peptic ulcer and inflammatory bowel disease (e.g., ulcerative colitis, chronic colitis, etc.); pancreatic diseases such as pancreatitis; hepatic diseases such as hepatitis and liver cirrhosis; diseases of bone or joint such as synovitis, arthritis, osteoarthritis, osteoporosis; autoimmune diseases such as rheumatoid arthritis, systemic lupus erythematosus and multiple sclerosis; dermal diseases such as dermatitis and eczema; cancer such as solid tumors and metastasis; rejection by organ transplantation; shock (e.g., septic shock, etc.); sepsis-induced systemic inflammatory response syndrome; and sexual dysfunction such as male sexual dysfunction (e.g., erectile dysfunction) and female sexual dysfunction (e.g., orgasmic dysfunction related to clitoral disturbances) in human being and animals.
The object peptide compounds of the present invention are novel and can be represented by the following general formula (I): 
wherein
R1 is benzofuranyl substituted by halogen, or styryl substituted by halogen,
R2 is phenyl, pyridyl, thienyl or thiazolyl, each of which is optionally substituted by one or more substituents selected from the group consisting of lower alkyl, lower alkoxy, halogen and trihalo(lower)alkyl, or lower alkyl optionally substituted by one or more halogen atoms,
R3 is hydrogen or lower alkoxy,
R4 and R5 are the same or different and each is hydrogen, lower alkyl or optionally protected hydroxy(lower)alkyl, and
R6 is hydrogen or lower alkyl.
Suitable pharmaceutically acceptable salts of the object compound (I) are conventional non-toxic salts and include, for example, a salt with a base or an acid addition salt such as a salt with an inorganic base, for example, an alkali metal salt (e.g., sodium salt, potassium salt, etc.), an alkaline earth metal salt (e.g., calcium salt, magnesium salt, etc.), an ammonium salt; a salt with an organic base, for example, an organic amine salt (e.g., triethylamine salt, pyridine salt, picoline salt, ethanolamine salt, triethanolamine salt, dicyclohexylamine salt, N,Nxe2x80x2-dibenzylethylenediamine salt, etc.); an inorganic acid addition salt (e.g., hydrochloride, hydrobromide, sulfate, phosphate, etc.); an organic carboxylic or sulfonic acid addition salt (e.g., formate, acetate, trifluoroacetate, maleate, tartrate, citrate, fumarate, methanesulfonate, benzenesulfonate, toluenesulfonate, etc.); and a salt with a basic or acidic amino acid (e.g., arginine, aspartic acid, gultamic acid, etc.).
In the above and subsequent descriptions of the present specification, suitable examples and illustration of the various definitions which the present invention intends to include within the scope thereof are explained in detail as follows.
The term xe2x80x9clowerxe2x80x9d is used to intend a group having 1 to 6, preferably 1 to 4, carbon atom(s), unless otherwise provided.
Suitable xe2x80x9chalogenxe2x80x9d includes, for example, fluorine, bromine, chlorine and iodine.
xe2x80x9cStyryl substituted by halogenxe2x80x9d means styryl which has halogen atom as a substituent on the benzene ring. Suitable examples of xe2x80x9cstyryl substituted by halogenxe2x80x9d include 2-(2-chlorophenyl)ethenyl, 2-(3-chlorophenyl)ethenyl, 2-(4-chlorophenyl)ethenyl, 2-(2-bromophenyl)ethenyl, 2-(3-bromophenyl)ethenyl, 2-(4-bromophenyl)ethenyl, 2-(2-fluorophenyl)ethenyl, 2-(3-fluorophenyl)ethenyl, 2-(4-fluorophenyl)ethenyl, and the like.
Suitable xe2x80x9clower alkylxe2x80x9d and xe2x80x9clower alkylxe2x80x9d moiety in the terms xe2x80x9ctrihalo(lower)alkylxe2x80x9d and xe2x80x9chydroxy(lower)alkylxe2x80x9d include straight or branched alkyl having 1 to 6 carbon atom(s), such as methyl, ethyl, propyl, isopropyl, butyl, isobutyl, sec-butyl, tert-butyl, pentyl, tert-pentyl and hexyl, in which more preferred one is C1-C4 alkyl.
Suitable xe2x80x9clower alkoxyxe2x80x9d includes straight or branched alkoxy having 1 to 6 carbon atom(s), such as methoxy, ethoxy, propoxy, isopropoxy, butoxy, isobutoxy, sec-butoxy, tert-butoxy, pentyloxy, tert-pentyloxy and hexyloxy, in which more preferred one is C1-C4 alkoxy.
Suitable examples of xe2x80x9ctrihalo(lower)alkylxe2x80x9d include trifluoromethyl, trichloromethyl, tribromomethyl, 2,2,2-trifluoroethyl and 3,3,3-trifluoropropyl, in which more preferred one is trifluoromethyl.
xe2x80x9cLower alkyl optionally substituted by one or more halogen atomsxe2x80x9d includes lower alkyl and halo(lower)alkyl. Suitable xe2x80x9chalo(lower)alkylxe2x80x9d includes lower alkyl substituted by 1 to 3 halogen atoms such as trifluoromethyl, trichloromethyl, tribromomethyl, 2,2,2-trifluoroethyl and 3,3,3-trifluoropropyl, in which more preferred one is trihalo(lower)alkyl.
xe2x80x9cOptionally protected hydroxy(lower)alkylxe2x80x9d includes hydroxy(lower)alkyl and protected hydroxy(lower)alkyl. Suitable examples of hydroxy-protecting group include lower alkyl (e.g., methyl, ethyl, propyl, isopropyl, butyl, sec-butyl, isobutyl, tert-butyl, pentyl, neopentyl, hexyl, etc.), acyl such as lower alkanoyl optionally substituted by one to three halogen atoms (e.g., formyl, acetyl, propanoyl, butanoyl, 2-methylpropanoyl, pentanoyl, 2,2-dimethylpropanoyl, hexanoyl, trichloroacetyl, trifluoroacetyl, etc.), optionally substituted phenyl(lower)alkyl, for example, mono(or di or tri)phenyl(lower)alkyl optionally substituted by nitro or lower alkoxy (e.g., benzyl, 4-nitrobenzyl, 4-methoxybenzyl, benzhydryl, trityl, etc.), trisubstituted silyl [e.g., tri(lower)alkylsilyl (e.g., trimethylsilyl, tert-butyldimethylsilyl, etc.), etc.], tetrahydropyranyl and the like.
Suitable xe2x80x9chydroxy(lower)alkylxe2x80x9d includes hydroxy(C1-C6)alkyl such as hydroxymethyl, 1-hydroxyethyl, 2-hydroxyethyl, 1-hydroxypropyl, 2-hydroxypropyl, 3-hydroxypropyl, 4-hydroxybutyl, 5-hydroxypentyl and 6-hydroxyhexyl.
Suitable xe2x80x9cprotected hydroxy(lower)alkylxe2x80x9d includes lower alkoxy(lower)alkyl such as (C1-C6)alkoxy(C1-C6)alkyl, for example, methoxymethyl, 1-methoxyethyl, 2-methoxyethyl, 1-methoxypropyl, 2-methoxypropyl, 3-methoxypropyl, ethoxymethyl, 2-ethoxyethyl, 3-ethoxypropyl and propoxymethyl.
Phenyl, pyridyl, thienyl or thiazolyl at R2 is optionally substituted by one or more, preferably one to three, substituents selected from the group consisting of lower alkyl, lower alkoxy, halogen and trihalo(lower)alkyl.
Suitable xe2x80x9camino-protecting groupxe2x80x9d includes, for example, acyl and conventional protecting group such as mono(or di or tri)aryl(lower)alkyl, for example, mono(or di or tri)phenyl (lower)alkyl (e.g., benzyl, trityl, etc.). Suitable examples of said acyl include aliphatic acyl such as lower alkanoyl which may be substituted by one to three halogen atoms (e.g., formyl, acetyl, propanoyl, butanoyl, 2-methylpropanoyl, pentanoyl, 2,2-dimethylpropanoyl, hexanoyl, trichloroacetyl, trifluoroacetyl, etc.), lower alkoxycarbonyl (e.g., methoxycarbonyl, ethoxycarbonyl, tert-butoxycarbonyl, tert-pentyloxycarbonyl, etc.), aryl(lower)alkoxycarbonyl [e.g., phenyl(lower)-alkoxycarbonyl (e.g., benzyloxycarbonyl, etc.), etc.] and the like.
Suitable xe2x80x9ccarboxy-protecting groupxe2x80x9d includes, for example, lower alkyl (e.g., methyl, ethyl, propyl, isopropyl, butyl, sec-butyl, isobutyl, tert-butyl, pentyl, neopentyl, hexyl, etc.), optionally substituted phenyl(lower)alkyl, for example, mono(or di or tri)phenyl(lower)alkyl which may be substituted by nitro (e.g., benzyl, 4-nitrobenzyl, benzhydryl, trityl, etc.) and the like.
The object compound (I) of the present invention can be prepared by the following processes.
Process (1) 
Process (2) 
wherein R1, R2, R3, R4, R5 and R6 are each as defined above.
The starting compounds can be prepared by the following processes.
Process (A) 
Process (B) 
Process (C) 
Process (D) 
Process (E) 
Process (F) 
Process (G) 
Process (H) 
Process (I) 
Process (J) 
Process (K) 
wherein R1, R2, R3, R1, R5 and R6 are each as defined above,
R7 and R8 are each amino-protecting group, and
R9 and R10 are each carboxy-protecting group.
The processes for preparing the object compound are explained in detail in the following.
Process (1)
The compound (I) or a salt thereof can be prepared by reacting the compound (II) or its reactive derivative at the carboxy group, or a salt thereof with the compound (III) or its reactive derivative at the amino group, or a salt thereof.
Suitable reactive derivative of the compound (III) includes Schiff""s base type imino or its tautomeric enamine type isomer formed by the reaction of the compound (III) with a carbonyl compound such as aldehyde, ketone or the like; a silyl derivative formed by the reaction of the compound (III) with a silyl compound such as N,O-bis(trimethylsilyl)acetamide, N-trimethylsilylacetamide or the like; a derivative formed by the reaction of the compound (III) with phosphorus trichloride or phosgene.
Suitable reactive derivative of the compound (II) includes an acid halide, an acid anhydride and an activated ester. The suitable example may be an acid chloride; an acid azide; a mixed acid anhydride with an acid such as substituted phosphoric acid (e.g., dialkylphosphoric acid, phenylphosphoric acid, diphenylphosphoric acid, dibenzylphosphoric acid, halogenated phosphoric acid, etc.), dialkylphosphorous acid, sulfurous acid, thiosulfuric acid, alkanesulfonic acid (e.g., methanesulfonic acid, ethanesulfonic acid, etc.), sulfuric acid, alkylcarbonic acid, aliphatic carboxylic acid (e.g., pivalic acid, pentanoic acid, isopentanoic acid, 2-ethylbutyric acid, trichloroacetic acid, etc.), and aromatic carboxylic acid (e.g., benzoic acid, etc.); a symmetrical acid anhydride; an activated amide with imidazole, 4-substituted imidazole, dimethylpyrazole, triazole or tetrazole; an activated ester (e.g., cyanomethyl ester, methoxymethyl ester, dimethyliminomethyl [(CH3)2N+=CHxe2x80x94] ester, vinyl ester, propargyl ester, p-nitrophenyl ester, 2,4-dinitrophenyl ester, trichlorophenyl ester, pentachlorophenyl ester, mesylphenyl ester, phenylazophenyl ester, phenyl thioester, p-nitrophenyl thioester, p-cresyl thioester, carboxymethyl thioester, pyranyl ester, pyridyl ester, piperidyl ester, 8-quinolyl thioester, etc.); or an ester with an N-hydroxy compound (e.g., N,N-dimethylhydroxylamine, 1-hydroxy-2-(1H)-pyridone, N-hydroxysuccinimide, N-hydroxybenzotriazole, N-hydroxyphthalimide, 1-hydroxy-6-chloro-1H-benzotriazole, etc.). These reactive derivatives can optionally be selected from them according to the kind of the compound (II) to be used.
The reaction is usually carried out in a conventional solvent such as water, acetone, dioxane, acetonitrile, chloroform, methylene chloride, ethylene chloride, tetrahydrofuran, ethyl acetate, N,N-dimethylformamide, pyridine or any other organic solvents which do not adversely affect the reaction, or the mixture thereof.
When the compound (II) is used in free acid form or its salt form in the reaction, the reaction is preferably carried out in the presence of a conventional condensing agent such as N,Nxe2x80x2-dicyclohexylcarbodiimide; N-cyclohexyl-Nxe2x80x2-morpholinoethyl-carbodiimide; N-cyclohexyl-Nxe2x80x2-(4-diethylaminocyclohexyl)-carbodiimide; N,Nxe2x80x2-diisopropylcarbodiimide; N-ethyl-Nxe2x80x2-(3-dimethylaminopropyl)carbodiimide; N,N-carbonyl-bis-(2-methylimidazole); pentamethyleneketene-N-cyclohexylimine; diphenylketene-N-cyclohexylimine; ethoxyacetylene; 1-alkoxy-1-chloroethylene; trialkyl phosphite; isopropyl polyphosphate; phosphorus oxychloride (phosphoryl chloride); phosphorus trichloride; thionyl chloride; oxalyl chloride; triphenylphosphine; 2-ethyl-7-hydroxybenzisoxazolium salt; 2-ethyl-5-(m-sulfophenyl)isoxazolium hydroxide intramolecular salt; 1-(p-chlorobenzenesulfonyloxy)-6-chloro-1H-benzotriazole; so-called Vilsmeier reagent prepared by the reaction of N,N-dimethylformamride with thionyl chloride, phosgene, phosphorus oxychloride, etc.; or the like.
The reaction may also be carried out in the presence of an organic or inorganic base such as an alkali metal bicarbonate, tri(lower)alkylamine, pyridine, N-(lower)alkylmorpholine, N,N-di(lower)alkylbenzylamine, or the like.
The reaction temperature is not critical, and the reaction is usually carried out under cooling to heating.
Process (2)
The compound (I) or a salt thereof can be prepared by reacting the compound (IV) or its reactive derivative at the carboxy group, or a salt thereof with the compound (V) or a salt thereof.
This reaction can be carried out in a similar manner to the reaction in the aforementioned Process (1), and therefore the reagents to be used and reaction conditions (e.g., solvent, reaction temperature, etc.) can be referred to those of Process (1).
Process (A)
The compound (VII) or a salt thereof can be prepared by reacting the compound (VI) or its reactive derivative at the carboxy group, or a salt thereof with the compound (V) or a salt thereof.
This reaction can be carried out in a similar manner to the reaction in the aforementioned Process (1), and therefore the reagents to be used and reaction conditions (e.g., solvent, reaction temperature, etc.) can be referred to those of Process (1).
Process (B)
The compound (VIII) or a salt thereof can be prepared by subjecting the compound (VII) or a salt thereof to elimination reaction of the amino-protecting group.
Suitable method of this elimination reaction includes conventional one such as hydrolysis, reduction and the like.
(i) For Hydrolysis:
The hydrolysis is preferably carried out in the presence of a base or an acid including Lewis acid.
Suitable base includes an inorganic base and an organic base such as an alkali metal (e.g., sodium, potassium, etc.), an alkaline earth metal (e.g., magnesium, calcium, etc.), the hydroxide or carbonate or hydrogencarbonate thereof, trialkylamine (e.g., trimethylamine, triethylamine, etc.), picoline, 1,5-diazabicyclo[4.3.0]non-5-one, or the like.
Suitable acid includes an organic acid (e.g., formic acid, acetic acid, propionic acid, trichloroacetic acid, trifluoroacetic acid, etc.), and an inorganic acid (e.g., hydrochloric acid, hydrobromic acid, sulfuric acid, hydrogen chloride, hydrogen bromide, etc.).
The elimination using. Lewis acid such as trihaloacetic acid (e.g., trichloroacetic acid, trifluoroacetic acid, etc.), or the like is preferably carried out in the presence of cation trapping agents (e.g., anisole, phenol, etc.). This reaction is usually carried out without solvent.
The reaction may be carried out in a conventional solvent such as water, alcohol (e.g., methanol, ethanol, isopropyl alcohol, etc.), tetrahydrofuran, dioxane, toluene, methylene chloride, ethylene dichloride, chloroform, N,N-dimethylformamide, N,N-dimethylacetamide or any other organic solvents which do not adversely affect the reaction, or a mixture thereof.
The reaction temperature is not critical and the reaction is usually carried out under cooling to warming.
(ii) For Reduction:
Reduction is carried out in a conventional manner, including chemical reduction and catalytic reduction.
Suitable reducing reagent to be used in chemical reduction are hydrides (e.g., hydrogen iodide, hydrogen sulfide, lithium aluminum hydride, sodium borohydride, sodium cyanoborohydride, etc.), or a combination of a metal (e.g., tin, zinc, iron, etc.) or metallic compound (e.g., chromium chloride, chromium acetate, etc.) and an organic acid or inorganic acid (e.g., formic acid, acetic acid, propionic acid, trifluoroacetic acid, p-toluenesulfonic acid, hydrochloric acid, hydrobromic acid, etc.).
Suitable catalysts to be used in catalytic reduction are conventional ones such as platinum catalysts (e.g., platinum plate, spongy platinum, platinum black, colloidal platinum, platinum oxide, platinum wire, etc.), palladium catalysts (e.g., spongy palladium, palladium black, palladium oxide, palladium on carbon, palladium hydroxide on carbon, colloidal palladium, palladium on barium sulfate, palladium on barium carbonate, etc.), nickel catalysts (e.g., reduced nickel, nickel oxide, Raney nickel, etc.), cobalt catalysts (e.g., reduced cobalt, Raney cobalt, etc.), iron catalysts (e.g., reduced iron, Raney iron, Ullman iron, etc.), and the like.
The reduction is usually carried out in a conventional solvent such as water, alcohol (e.g., methanol, ethanol, isopropyl alcohol, etc.), tetrahydrofuran, dioxane, toluene, methylene chloride, ethylene dichloride, chloroform, N,N-dimethylformamide, N,N-dimethylacetamide or any other organic solvents which do not adversely affect the reaction, or a mixture thereof.
Additionally, in case that the above-mentioned acids to be used in chemical reduction are in a liquid state, they can also be used as a solvent.
The reaction temperature of this reduction is not critical and the reaction is usually carried out under cooling to warming.
Process (C)
The compound (X) or a salt thereof can be prepared by reacting the compound (IX) or its reactive derivative at the carboxy group, or a salt thereof with the compound (VIII) or its reactive derivative at the amino group, or a salt thereof.
This reaction can be carried out in a similar manner to the reaction in the aforementioned Process (1), and therefore the reagents to be used and reaction conditions (e.g., solvent, reaction temperature, etc.) can be referred to those of Process (1).
Process (D)
The compound (III) or a salt thereof can be prepared by subjecting the compound (X) or a salt thereof to elimination reaction of the amino-protecting group.
This reaction can be carried out in a similar manner to the reaction in the aforementioned Process (B), and therefore the reagents to be used and reaction conditions (e.g., solvent, reaction temperature, etc.) can be referred to those of Process (B).
Process (E)
The compound (XII) or a salt thereof can be prepared by reacting the compound (II) or its reactive derivative at the carboxy group, or a salt thereof with the compound (XI) or its reactive derivative at the amino group, or a salt thereof.
This reaction can be carried out in a similar manner to the reaction in the aforementioned Process (1), and therefore the reagents to be used and reaction conditions (e.g., solvent, reaction temperature, etc.) can be referred to those of process (1).
Process (F)
The compound (XIII) or a salt thereof can be prepared by subjecting the compound (XII) or a salt thereof to elimination reaction of the carboxy-protecting group.
This reaction can be carried out in a similar manner to the reaction in the aforementioned Process (B), and therefore the reagents to be used and reaction conditions (e.g., solvent, reaction temperature, etc.) can be referred to those of Process (B).
Process (G)
The compound (XV) or a salt thereof can be prepared by reacting the compound (XIII) or its reactive derivative at the carboxy group, or a salt thereof with the compound (XIV) or its reactive derivative at the amino group, or a salt thereof.
This reaction can be carried out in a similar manner to the reaction in the aforementioned Process (1), and therefore the reagents to be used and reaction conditions (e.g., solvent, reaction temperature, etc.) can be referred to those of Process (1).
Process (H)
The compound (IV) or a salt thereof can be prepared by subjecting the compound (XV) or a salt thereof to elimination reaction of the carboxy-protecting group.
This reaction can be carried out in a similar manner to the reaction in the aforementioned Process (B), and therefore the reagents to be used and reaction conditions (e.g., solvent, reaction temperature, etc.) can be referred to those of Process (B).
Process (I)
The compound (XVI) or a salt thereof can be prepared by reacting the compound (IX) or its reactive derivative at the carboxy group, or a salt thereof with the compound (XIV) or its reactive derivative at the amino group, or a salt thereof.
This reaction can be carried out in a similar manner to the reaction in the aforementioned Process (1), and therefore the reagents to be used and reaction conditions (e.g., solvent, reaction temperature, etc.) can be referred to those of Process (1).
Process (J)
The compound (XVII) or a salt thereof can be prepared by subjecting the compound (XVI) or a salt thereof to elimination reaction of the amino-protecting group.
This reaction can be carried out in a similar manner to the reaction in the aforementioned Process (B), and therefore the reagents to be used and reaction conditions (e.g., solvent, reaction temperature, etc.) can be referred to those of Process (B).
Process (K)
The compound (XV) or a salt thereof can be prepared by reacting the compound (II) or its reactive derivative at the carboxy group, or a salt thereof with the compound (XVII) or its reactive derivative at the amino group, or a salt thereof.
This reaction can be carried out in a similar manner to the reaction in the aforementioned Process (1), and therefore the reagents to be used and reaction conditions (e.g., solvent, reaction temperature, etc.) can be referred to those of Process (1).
Suitable salts of the starting compounds and their reactive derivatives in Processes (1) and (2) and Processes (A) to (K) can be referred to the ones as exemplified for the compound (I).
The compounds obtained by the above process can be isolated and purified by a conventional method such as pulverization, recrystallization, column chromatography, reprecipitation, or the like.
It is to be noted that the compound (I) and the other compounds may include one or more stereoisomer(s) such as optical isomer(s) and geometrical isomer(s) due to asymmetric carbon atom(s) and double bond(s), and all of such isomers and mixtures thereof are included within the scope of this invention.
The object compounds (I) and pharmaceutically acceptable salts thereof include solvates [e.g., enclosure compounds (e.g., hydrate, etc.)].
The object compounds (I) and pharmaceutically acceptable salts thereof possess a strong inhibitory activity on the production of nitric oxide (NO).
Accordingly, the object compounds (I) and pharmaceutically acceptable salts thereof are expected to possess a nitric oxide synthase (NOS)-inhibitory activity or a NOS-production inhibitory activity.
Accordingly, the object compounds (I) and pharmaceutically acceptable salts thereof are useful for prevention and/or treatment of NO-mediated diseases in human being and animals, including respiratory diseases such as adult respiratory distress syndrome (ARDS) and asthma; cardiovascular diseases such as cardiovascular ischemia, myocarditis, heart failure, hypotension and atherosclerosis; endocrine diseases such as diabetes (e.g., insulin-dependent diabetes mellitus, etc.), complications of diabetes mellitus (e.g., diabetic nephropathy, diabetic retinopathy, diabetic neuropathy, etc.) and gout; renal diseases such as glomerulonephritis and renal failure; gastrointestinal diseases such as peptic ulcer and inflammatory bowel disease (e.g., ulcerative colitis, chronic colitis, etc.); pancreatic diseases such as pancreatitis; hepatic diseases such as hepatitis and liver cirrhosis; diseases of bone or joint such as synovitis, arthritis, osteoarthritis, osteoporosis; autoimmune diseases such as rheumatoid arthritis, systemic lupus erythematosus and multiple sclerosis; dermal diseases such as dermatitis and eczema; cancer such as solid tumors and metastasis; rejection by organ transplantation; shock (e.g., septic shock, etc.); and sepsis-induced systemic inflammatory response syndrome.
The object compounds (I) and pharmaceutically acceptable salts thereof are also useful for prevention and/or treatment of No-mediated nervous diseases including central nervous system diseases such as CNS disorders, cerebrovascular diseases (e.g., cerebral infarction, cerebral ischemia, cerebral hemorrhage, etc.), migraine, Alzheimer""s disease; peripheral nervous system diseases such as neuritis, pain (e.g., postherpetic neuralgia, reflex sympathetic dystrophy (RSD), causalgia, deafferentation pain syndrome, neuropathic pain, etc.), allodynia, hyperalgesia, neurological disorders and neuroprotection; Parkinson""s disease; and amyotrophic lateral sclerosis.
Additionally, the object compounds (I) and pharmaceutically acceptable salts thereof are useful for treatment of sexual dysfunction such as male sexual dysfunction including erectile dysfunction, and female sexual dysfunction including orgasmic dysfunction related to clitoral disturbances.
Further, the object compounds (I) and pharmaceutically acceptable salts thereof are useful for prevention and/or treatment of NO-mediated ophthalmic diseases, including conjunctive diseases such as conjunctivitis (e.g., allergic conjunctivitis, vernal conjunctivitis, keratoconjunctivitis sicca, viral conjunctivitis, bacterial conjunctivitis, etc.); uveal diseases such as uveitis (e.g., Behcet disease, Harada disease, sympathetic ophthalmia, sarcoidosis, diabetic iritis, etc.); scleral diseases such as scleritis; corneal diseases such as corneal neovascularization, keratitis, corneal edema, corneal opacity, corneal dystrophy, keratoconus and neuroparalytic keratitis; retinal, vitreous diseases such as diabetic retinopathy, retinal artery occlusion, retinal vein occlusion, central serous chorioretinopathy, central hemorrhagic chorioretinitis, macular degeneration (e.g., age-related macular degeneration, etc.), retinal detachment, retinal pigmentary degeneration, macular neovascularization, macular hole, proliferative vitreoretinopathy, vitreous hemorrhage and vitreous opacity; lens diseases such as cataract (e.g., senile cataract, traumatic cataract, diabetic cataract, atopic cataract, etc.); glaucoma such as primary open-angle glaucoma, primary angle-closure glaucoma, normal tension glaucoma and neovascular glaucoma; ocular hypertension; vision disorders such as amblyopia, color vision defect and night blindness; refractive errors such as astigmatism, hyperopia, myopia and presbyopia; and lacrimal apparatus diseases such as dry eye syndromes, lacrimal duct obstruction and dacryocystitis.
In order to illustrate the usefulness of the object compound (I), the pharmacological test result of the compound (I) is shown in the following.
Test Compound:
Compound (a): 5-Chloro-N-[(1S)-2-[[2-[4-(4-chlorophenyl)-1-piperazinyl]-2-oxoethyl]amino]-2-oxo-1-(2-pyridylmethyl)ethyl]-1-benzofuran-2-carboxamide
Compound (b): (2E)-3-(4-Chlorophenyl)-N-[(1S)-2-[[2-((2S)-2-methyl-4-phenyl-1-piperazinyl)-2-oxoethyl]amino]-2-oxo-1-(2-pyridylmethyl)ethyl]-2-propenamide
Compound (c): (2E)-3-(4-Chlorophenyl)-N-[(1S)-2-[[2-[(2R,6S)-2,6-dimethyl-4-(4-methylphenyl)-1-piperazinyl]-2-oxoethyl]amino]-2-oxo-1-(2-pyridylmethyl)ethyl]-2-propenamide
Compound (d): (2E)-3-(4-Chlorophenyl)-N-[(1S)-2-oxo-2-[[2-oxo-2-[4-(3,3,3-trifluoropropyl)-1-piperazinyl]ethyl]amino]-1-(2-pyridylmethyl) ethyl]-2-propenamide
Compound (e): (2E)-3-(4-Chlorophenyl)-N-[(1S)-2-oxo-2-[[2-oxo-2-[4-(4-trifluoromethylphenyl)-1-piperazinyl]ethyl]amino]-1-(2-pyridylmethyl)ethyl]-2-propenamide
Compound (f): 5-Chloro-N-[(1S)-2-[[2-[4-(4-methoxyphenyl)-1-piperazinyl]-2-oxoethyl]amino]-2-oxo-1-(2-pyridylmethyl) ethyl]-1-benzofuran-2-carboxamide
Test 1: Assay for Inhibitory Activity on the Production of Nitric Oxide
The murine macrophage cell line RAW264.7 (American Type Culture Collection, No. TIB71) was used in this study. RAW264.7 cells were grown on F75 plastic culture flasks at 37xc2x0 C., 5% in Dulbecco""s modified Eagle""s medium (DMEM) supplemented with L-glutamine, penicillin, streptomycin and 10% heat-inactivated fetal bovine serum. They were removed from culture flasks by rubber cell scraper and were centrifuged and resuspended in DMEM without phenol red. They were plated in 96-well microtiter plates (105 cells per well) and allowed to adhere over 2 hours. The test samples were added and the cells were preincubated for 1 hour. Thereafter the cells were activated with both of lipopolysaccharide (LPS) (1 xcexcg/ml) and interferon xcex3 (INF xcex3) (3 u/ml) for 18-24 hours. An equal volume of Griess reagent (1% sulfanilamide/0.1% N-naphthylethylenediamine dihydrochloride/2.5% H3PO4) was added and the cells were incubated at room temperature for 10 minutes. The absorbance was read at 570 nm using microplate reader and NO2xe2x88x92 was measured using NaNO2 as a standard.
Test Result:
Test 2: Protective Effect of the Compound (I) Combined with FK506 on Rat Cardiac Allograft
Method:
Experiments were performed on male Lewis and ACI rats weighing 175-200 g. Rats were anesthetized with sodium pentobarbital (50 mg/kg, i.p.), and underwent allogeneic (Lewis donor to ACI recipient) heterotopic cardiac transplantation. Experimental groups were divided into single-drug group and combined-drug group. Single-drug dose of FK506, which was prepared in a manner similar to that disclosed in EP-0184162, was 0.32 mg/kg. Combined-drug dose was FK506 (0.32 mg/kg)+the compound (I) (10 mg/kg). The grafted hearts were monitored by daily palpation where complete rejection was defined as the cessation of palpable contractile activity. Each drug was suspended in a solution of 0.5% methylcellulose, and administered by daily gastric intubation in a volume of 5 ml/kg of body weight for 14 days.
The combination of the compound (I) and FK506 dramatically prolonged the graft survival.
The above experimental results indicate that the activity and/or efficacy of an immunosuppressant in rejection of transplantation can be remarkably and synergistically increased by administering compound (I) in combination, which has a strong inhibitory activity on the production of nitric oxide.
For therapeutic administration, the object compound (I) of the present invention and pharmaceutically acceptable salts thereof are used in the form of a conventional pharmaceutical preparation in admixture with a conventional pharmaceutically acceptable carrier such as an organic or inorganic solid or liquid excipient which is suitable for oral, parenteral or external administration. The pharmaceutical preparation may be compounded in a solid form such as granule, capsule, tablet, dragee, suppository or ointment, or in a liquid form such as solution, suspension or emulsion for injection, intravenous drip, ingestion, eye drop, etc. If needed, there may be included in the above preparation auxiliary substance such as stabilizing agent, wetting or emulsifying agent, buffer or any other commonly used additives.
The effective ingredient may usually be administered in a unit dose of 0.001 mg/kg to 500 mg/kg, preferably 0.01 mg/kg to 10 mg/kg, 1 to 4 times a day. However, the above dosage may be increased or decreased according to age, body weight and conditions of the patient or administering method.
According to the present invention, a pharmaceutical composition comprising FK506 and the compound (I) or a pharmaceutically acceptable salt thereof is provided. A pharmaceutical composition comprising FK506 and the compound (I) or a pharmaceutically acceptable salt thereof is useful as an immunosuppressant. For example, the pharmaceutical composition of the present invention is useful for the prevention or treatment of rejection by organ transplantation.
When the compound (I) is used in combination with FK506, a ratio by weight of the compound (I) or a pharmaceutically acceptable salt thereof to FK506 is in the range of 0.1/1-1000/1, preferably in the range of 1/1-100/1.
The preferred embodiments of the peptide compound of the present invention represented by the general formula (I) are as follows.
1) The compound of the formula (I) wherein R1 is benzofuranyl substituted by halogen.
2) The compound of the formula (I) wherein R1 is styryl substituted by halogen.
3) The compound of the formula (I) wherein R2 is phenyl, pyridyl, thienyl or thiazolyl, each of which is optionally substituted by one or more substituents selected from the group consisting of lower alkyl, lower alkoxy, halogen and trihalo(lower)alkyl, or trihalo(lower)alkyl.
4) The compound of the formula (I) wherein R2 is phenyl optionally substituted by one or more substituents selected from the group consisting of lower alkyl, lower alkoxy, halogen and trihalo(lower)alkyl, or trihalo(lower)alkyl.
5) The compound of the formula (I) wherein R2 is pyridyl, thienyl or thiazolyl, each of which is optionally substituted by one or more substituents selected from the group consisting of halogen and trihalo(lower)alkyl.
6) The compound of the formula (I) wherein R4 and R5 are the same or different and each is hydrogen, lower alkyl, hydroxy(lower)-alkyl or lower alkoxy(lower)alkyl.
The following Preparations and Examples are given for the purpose of illustrating the present invention in detail.
Preparation 1
tert-Butoxy-N-[2-[4-(4-chlorophenyl)-1-piperazinyl]-2-oxoethyl]formamide
To a suspension of 2-(tert-butoxycarbonylamino)acetic acid (16.3 g) and 1-(4-chlorophenyl)piperazine hydrochloride (21.7 g) in N,N-dimethylformamide (200 ml) were added 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride (19.6 g) and N-hydroxybenzotriazole (13.8 g) at 0xc2x0 C. To this suspension was added dropwise triethylamine (10.4 g). The mixture was stirred at room temperature for 20 hours. The reaction mixture was poured into ice-cold water and the resulting solid was collected by filtration. The solid was washed successively with saturated aqueous sodium hydrogencarbonate solution and water. The solid was dried to give the title compound (29 g).
1H-NMR (300 MHz, CDCl3) xcex4 3.09-3.18 (m, 4H), 3.51-3.59 (m, 2H), 3.74-3.82 (m, 2H), 4.01 (d, J=5 Hz, 2H), 5.50 (m, 1H), 6.84 (d, J=8 Hz, 2H), 7.23 (d, J=8 Hz, 2H).
Preparation 2
2-Amino-1-[4-(4-chlorophenyl)-1-piperazinyl]ethan-1-one dihydrochloride
To a solution of tert-butoxy-N-[2-[4-(4-chlorophenyl)-1-piperazinyl]-2-oxoethyl]formamide (31 g) in methanol (60 ml) was added 4N hydrogen chloride in ethyl acetate (219 ml) at 0xc2x0 C. and the mixture was stirred at 0xc2x0 C. for 30 minutes. The reaction mixture was allowed to warm to room temperature, stirred for 1 hour and diluted with ethyl acetate. The resulting solid was collected by filtration and washed with ethyl acetate. The solid was dried to give the title compound (29 g).
1H-NMR (300 MHz, DMSO-d6) xcex4 3.11-3.28 (m, 4H), 3.55 (m, 2H), 3.66 (m, 2H), 3.92 (m, 2H), 7.04 (d, J=9 Hz, 2H), 7.29 (d, J=9 Hz, 2H), 8.22 (br, 2H).
Preparation 3
(2S)-2-(tert-Butoxycarbonylamino)-N-[2-[4-(4-chlorophenyl)-1-piperazinyl]-2-oxoethyl]-3-(2-pyridyl)propanamide
Diphenylphosphoryl azide (25 g) was added to a solution of (2S)-2-(tert-butoxycarbonylamino)-3-(2-pyridyl)propanoic acid (23 g) in N,N-dimethylformamide (230 ml) at 5xc2x0 C. To the mixture was added a solid of 2-amino-1-[4-(4-chlorophenyl)-1-piperazinyl]-ethan-1-one dihydrochloride (28.2 g) at 5xc2x0 C. with stirring and then diisopropylethylamine (47 ml) was added dropwise at 7-10xc2x0 C. The mixture was stirred at 7-10xc2x0 C. for 30 minutes and at room temperature for 2 hours. The mixture was diluted with saturated aqueous sodium hydrogencarbonate solution and extracted with ethyl acetate. The aqueous layer was extracted with two 500-ml portions of ethyl acetate, and the combined organic layers were extracted with 1N hydrochloric acid (150 mlxc3x972). The combined aqueous layers were basified (pH 9) with sodium hydrogencarbonate and extracted with ethyl acetate. The organic layer was washed thoroughly with aqueous sodium hydrogencarbonate solution and brine, dried over magnesium sulfate and concentrated in vacuo to give the title compound (42 g).
1H-NMR (300 MHz, CDCl3) xcex4 1.44 (s, 9H), 3.11 (m, 4H), 3.23 (m, 1H), 3.34 (m, 1H), 3.53 (m, 2H), 3.75 (m, 2H), 4.04 (d, J=5 Hz, 2H), 4.64 (m, 1H), 6.38 (m, 1H), 6.83 (d, J=8 Hz, 2H), 7.10-7.27 (m, 4H), 7.59 (m, 1H), 7.77 (br, 1H), 8.52 (d, J=5 Hz, 1H).
Preparation 4
(2S)-2-Amino-N-[2-[4-(4-chlorophenyl)-1-piperazinyl]-2-oxoethyl]-3-(2-pyridyl)propanamide
To a solution of (2S)-2-(tert-butoxycarbonylamino)-N-[2-[4-(4-chlorophenyl)-1-piperazinyl]-2-oxoethyl]-3-(2-pyridyl)-propanamide (42 g) in methanol (84 ml) was added 4N hydrogen chloride in ethyl acetate (209 ml) at 0xc2x0 C. and the mixture was stirred at 0xc2x0 C. for 30 minutes. The reaction mixture was allowed to warm to room temperature, stirred for 2 hours and diluted with ethyl acetate. The resulting solid was collected by filtration and washed with ethyl acetate. The solid was poured into aqueous sodium hydrogencarbonate solution and the mixture was extracted with chloroform. The aqueous layer was extracted twice with chloroform. The combined organic layers were dried over sodium sulfate and concentrated in vacuo to give a crude product of the title compound (27 g).
1H-NMR (300 MHz, DMSO-d6) xcex4 2.74 (dd, J=13 Hz,9 Hz, 1H), 3.06-3.23 (m, 5H), 3.50-3.70 (m, 5H), 3.93-4.12 (m, 2H), 6.97 (d, J=9 Hz, 2H), 7.17-7.34 (m, 4H), 7.70 (m, 1H), 8.24 (m, 1H), 8.49 (m, 1H).